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Modelling Cellular Excitability Basic reference: Keener and Sneyd, Mathematical Physiology Cardiac Cell Nattel and Carlsson Nature Reviews Drug Discovery 5, 1034–1049 (December 2006) Neuromuscular Junction Neuron Cell membranes, ion channels and receptors Basic problem • The cell is full of stuff. Proteins, ions, fats, etc. • Ordinarily, these would cause huge osmotic pressures, sucking water into the cell. • The cell membrane has no structural strength, and the cell would burst. Basic solution • Cells carefully regulate their intracellular ionic concentrations, to ensure that no osmotic pressures arise • As a consequence, the major ions Na+, K+, Cl- and Ca2+ have different concentrations in the extracellular and intracellular environments. • And thus a voltage difference arises across the cell membrane. • Essentially two different kinds of cells: excitable and nonexcitable. • All cells have a resting membrane potential, but only excitable cells modulate it actively. Typical ionic concentrations (in mM) Squid Giant Axon Frog Sartorius Muscle Human Red Blood Cell Intracellular Na+ 50 13 19 K+ 397 138 136 Cl- 40 3 78 Na+ 437 110 155 K+ 20 2.5 5 Cl- 556 90 112 Extracellular The Nernst equation [S]i=[S’]i Vi [S]e=[S’]e Ve Permeable to S, but not S’; S and S’ have opposite charge RT [ S ]e Vi Ve ln zF [ S ]i (The Nernst potential) Equilibrium is reached when the electric field exactly balances the diffusion of S. In the case of a single ion species the net current is zero at the Nernst potential. However this is not true when more than one type of ions can cross the membrane. Note: equilibrium only. Tells us nothing about the current. In addition, there is very little actual ion transfer from side to side. Resting potential • No ions are at equilibrium, so there are continual background currents. At steady-state, the net current is zero, not the individual currents. • The pumps must work continually to maintain these concentration differences and the cell integrity. • The resting membrane potential depends on the model used for the ionic currents. gNa (V VNa ) gK (V VK ) 0 Vsteady gNaVNa gKVK gNa gK linear current model (long channel limit) VF + + 2 F 2 [Na + ]i [Na+ ]e exp( VF ) [K ] [K ] exp( ) F i e RT RT PNa V P V K 0 VF VF 1 exp( RT ) 1 exp( RT ) RT RT RT PNa [Na+ ]e PK [K + ]e Vsteady ln GHK current model F PNa [Na+ ]i PK [K + ]i (short channel limit) Electrical circuit model of cell membrane outside C Iionic How to model this is the crucial question! inside dV C Iionic 0 dt Vi Ve V C dV/dt Simplifications • In some cells (electrically excitable cells), the membrane potential is a far more complicated beast. • To simplify modelling of these types of cells, it is simplest just to assume that the internal and external ionic concentrations are constant. • Justification: Firstly, it takes only small currents to get large voltage deflections, and thus only small numbers of ions cross the membrane. Secondly, the pumps work continuously to maintain steady concentrations inside the cell. • So, in these simpler models the pump rate never appears explicitly, and all ionic concentrations are treated as known and fixed. Steady-state vs instantaneous I-V curves • So far we have discussed how the current through a single open channel depends in the membrane potential and the ionic concentrations on either side of the membrane. • But in a population of channels, the total current is a function of the single-channel current, and the number of open channels. • When V changes, both the single-channel current changes, as well as the proportion of open channels. But the first change happens almost instantaneously, while the second change is a lot slower. I g(V,t) (V ) Proportion of open channels I-V curve of single open channel Hodgkin Huxley Alan Lloyd Hodgkin and Andrew Huxley described the model in 1952 to explain the ionic mechanisms underlying the initiation and propagation of action potentials in the squid giant axon. They received the 1963 Nobel Prize in Physiology or Medicine for this work. Example: Na+ and K+ channels Experimental data: K+ conductance If voltage is stepped up and held fixed, gK increases to a new steady level. four subunits gK gK n dn (V )(1 n) (V )n dt 4 n (V ) rate of rise gives n dn n (V ) n dt time constant steady-state Now just fit to the data. steady state gives n∞ K+ channel gating If the channel consists of two identical subunits, each of which can be closed or open then: S00 S01 S0 S10 dx 0 x1 2x 0 dt dx 2 x1 2x 2 dt x 0 x1 x 2 1 2 S1 2 S11 x 0 (1 n) 2 x1 2n(1 n) x2 n 2 dn (1 n) n dt S2 Experimental data: Na+ conductance If voltage is stepped up and held fixed, gNa increases and then decreases. gNa gNa m h 3 Four subunits. Three switch on. One switches off. dh h (V ) h dt dm m (V ) m (V ) m dt h (V ) time constant steady-state Fit to the data is a little more complicated now, but still easy in principle. Na+ channel gating If the channel consists of multiple subunits of two different types, m and h, each of which can be closed or open then: S00 2 S01 S10 2 2 S11 S20 2 x 21 m 2 h Si j activation S21 inactivation the fraction of channels in state S21 dm (1 m) m dt dh (1 h) h dt activation inactivation Hodgkin-Huxley equations applied current dV gK n 4 (V VK ) gNa m 3 h(V VNa ) gL (V VL ) Iapp 0 dt dn n (V ) n (V ) n generic leak dt dm dh m (V ) m (V ) m, h (V ) h (V ) h dt dt C activation (increases with V) much smaller than the others inactivation (decreases with V) An action potential • gNa increases quickly, but then inactivation kicks in and it decreases again. • gK increases more slowly, and only decreases once the voltage has decreased. • The Na+ current is autocatalytic. An increase in V increases m, which increases the Na+ current, which increases V, etc. • Hence, the threshold for action potential initiation is where the inward Na+ current exactly balances the outward K+ current. The fast phase plane: I dV gK n 04 (V VK ) gNa m 3 h0 (V VNa ) gL (V VL ) Iapp 0 dt dm m (V ) m (V ) m dt C n and h are slow, and so stay approximately at their steady states while V and m change quickly The fast phase plane: II h0 decreasing n0 increasing As n and h change slowly, the dV/dt nullcline moves up, ve and vs merge in a saddle-node bifurcation, and disappear. vr is the only remaining steadystate, and so V returns to rest. In this analysis, we simplified the four-dimensional phase space by taking series of two-dimensional cross-sections, those with various fixed values of n and h. The fast-slow phase plane Take a different cross-section of the 4-d system, by setting m=m∞(v), and using the useful fact that n + h = 0.8 (approximately). Why? Who knows. It just is. Thus: dV C gK n 4 (V VK ) gNa m3 (0.8 n)(V VNa ) gL (V VL ) Iapp 0 dt dn n (V ) n (V ) n dt depolarization (Iapp >0) Oscillations When a current is applied across the cell membrane, the HH equations can exhibit oscillatory action potentials. C dV Iionic Iapplied 0 dt V HB HB Iapplied Where does it go from here? • More detailed models - Traub, Golomb, Purvis, … . • Simplified models - FHN, Morris Lecar, Hindmarsh-Rose… • Forced oscillations of single cells - APD alternans, Wenckebach patterns. • Other simplified models - Integrate and Fire, Poincare oscillator • Networks and spatial coupling (neuroscience, cardiology, …) Some further references • Koch (1999) Simplified models of single neurons. • Rinzel & Ermentrout (1997) Analysis of neural excitability and oscillation. • Gerstner & Kempter (2002) Spiking neural models • Koch (1999) Phase Space Analysis of Neural Excitability Why Simplified Models? • Analysis of the dynamical behaviors of single neurons • Reduce the computational load • To be used in network models From Compartmental models to Point Neurons Axon hillock (Soma) Point Neurons • General Form : dV p C g i xi i yiqi (Vi V ) I Synaptic dt i dx x (Vm ) x dt x (Vm ) I synaptic g si (t )(Vi V ) i Two Dimensional Neurons • Enables phase plane analysis • Most important variants – Fitzhugh-Nagumo Model – Morris-Lecar Model • Software: XPPAUT, MatCont, DDEBifTool FitzHugh-Nagumo model Figure 1: Circuit diagram of the tunnel-diode nerve model of Nagumo et al. (1962). Fast variable FitzHugh modified the Van der Pol equations for the nonlinear relaxation oscillator. The result had a stable resting state, from which it could be excited by a sufficiently large electrical stimulus to produce an impulse. A large enough constant current stimulus produced a train of impulses (FitzHugh 1961, 1969). Slow (recovery) variable http://www.scholarpedia.org/article/FitzHugh-Nagumo_model How do we analyze this class of models? • Phase plane: Study the dynamics in the (V,w)plane rather than V or w versus time • Nullclines: Determine the curves along which one ofthe time derivatives is 0 • Steady states: At the intersections of the two nullclines both derivatives are 0, so the system is at rest • Direction arrows: The nullclines divide up the plane, and the direction of flow in each region can be determined Fitzhugh-Nagumo Model (1) • A simplification of HH model C dV g Na m3h (VNa V ) g K n 4 (VK V ) g L (VL V ) I ext dt n dn n n dt m dm m m dt dh h h h dt m is much faster than the others: m m Fitzhugh-Nagumo Model (2) • Eliminating the fast dynamics C dV 3 g Na m h (VNa V ) g K n 4 (VK V ) g L (VL V ) I ext dt n dn n n dt h dh h h dt Blue: Full system Red: Reduced System Fitzhugh-Nagumo Model (3) • Eliminating h C dV 3 g Na m h (VNa V ) g K n 4 (VK V ) g L (VL V ) I ext dt n dn n n dt h dh h h dt h(t ) 0.8 n(t ) Fitzhugh-Nagumo Model (4) • Fitzhugh-Nagumo 2D model: C n dV 3 g Na m (0.8 n) (VNa V ) g K n 4 (VK V ) g L (VL V ) I ext dt dn n n dt dn dV dt dt V : Fast System n : Slow System Fitzhugh-Nagumo Model (5) • Nullclines: n 0 V <0 n nullcline n 0 V <0 V 0 Orbit or Spiral? V >0 n 0 n 0 V nullcline Fitzhugh-Nagumo Model (6) • Fitzhugh- Nagumo equations: dV V3 V W I dt 3 dW (V a bW ) dt 2 1.5 1 0.5 a 0 . 7 , b 0 .8 0.08 W 0 -0.5 -1 -1.5 -2 -3 -2 -1 0 1 2 V Qualitatively captures the properties of the exact model 3 Analysis of Fitzhugh-Nagumo System (1) dV V3 V W I dt 3 dW (V a bW ) dt • Jacobian: a 0 . 7 , b 0 .8 0.08 (1 V 2 ) A -1 -b Fixed point for I=0 : V=-1.20, W=0.625 Eigen values of A (l 2 + (V 2 -1+ bj )l + (V 2 -1)bj + j = 0) : l1,2 = -0.5 ± 0.42i The fixed point is: Stable Spiral Analysis of Fitzhugh-Nagumo System (2) • Response of the resting system (I=0) to a current pulse: 2 1.5 1 0.5 W 0 -0.5 -1 -1.5 -2 -3 -2 -1 0 V 1 2 3 Analysis of Fitzhugh-Nagumo System (3) • Response of the resting system (I=0) to a current pulse: 2 1.5 1 0.5 W 0 -0.5 -1 -1.5 -2 -3 -2 -1 0 1 2 3 V Threshold is due to fast sodium gating (V nullcline) Hyperpolarization and its termination is due to sodium/potassium channel Analysis of Fitzhugh-Nagumo System (4) • Response to a steady current : dV V3 V W I dt 3 dW (V a bW ) dt Jacobian: a 0 . 7 , b 0 .8 0.08 (1 V 2 ) A -1 -b Fixed point for I=1 : V=0.41, W=1.39 Eigen values of A (l 2 + (V 2 -1+ bj )l + (V 2 -1)bj + j = 0) : l1,2 = 0.41± 0.32i The fixed point is: Unstable spiral Analysis of Fitzhugh-Nagumo System (4) • Response of the resting system (I=1) to a steady current: 2 1.5 1 0.5 W 0 -0.5 -1 Stable Limit Cycle -1.5 -2 -3 -2 -1 0 V 1 2 3 Analysis of Fitzhugh-Nagumo System (4) • Response of the resting system (I=1) to a steady current: 2 1.5 1 0.5 W 0 -0.5 -1 -1.5 -2 -3 -2 -1 0 V 1 2 3 Bifurcation • By increasing the parameter – I the stable fix point renders unstable – A stable limit cycle appears • If by changing a parameter qualitative behavior of a system changes, this phenomenon is called bifurcation and the parameter is called bifurcation parameter Onset of oscillation with nonzero frequency • In the resting (I=0) the fixed point is a stable spiral The imaginary part of the eigenvalue is not zero and the real part is negative In the bifurcation, the fixed point loses its stability the real part of eigen value becomes positive and the imaginary part remains non-zero The frequency of oscillation is proportional to the magnitude of the imaginary part of the eigenvalue By increasing I, the oscillation onset starts with non-zero frequency Hopf Bifurcation IF response of Fitzhugh-Nagumo model 30 f 0 0 0.25 0.5 I 0.75 1 Neuron Type I / Type II • Gain functions of type I and Type II neurons I II Neural Coding Type I : Axon Hillock (Soma) of most neurons Type II: Axons of Most neurons, whole body of non-adaptive cortical interneurons, the spinal neurons FitzHugh-Nagumo model Figure 3: Absence of all-or-none spikes in the FitzHugh-Nagumo model. Figure 4: Excitation block in the FitzHughNagumo model. Fast variable Figure 2: Phase portrait and physiological state diagram of FitzHugh-Nagumo model (modified from FitzHugh 1961). Slow (recovery) variable http://www.scholarpedia.org/article/FitzHugh-Nagumo_model Question • What happens if we feed the FitzhughNagumo neuron with a strong inhibitory pulse? • Post inhibitory rebound spike W V FitzHugh-Nagumo model Figure 5: Anodal break excitation (postinhibitory rebound spike) in the FitzHughNagumo model. Fast variable Figure 6: Spike accommodation to slowly increasing stimulus in the FitzHughNagumo model. Slow (recovery) variable http://www.scholarpedia.org/article/FitzHugh-Nagumo_model Bursting Neurons • Adding another slow process (Eugene Izhikevich 2000) – Three dimensional phase plane Dynamic Clamp/Conductance Injection A method to assess how biophysically-defined ionic conductances shape the firing patterns of neurones Or A Physiologist’s dream: Adding and removing defined channel types without having to resort to pharmacology or molecular biology How would the cell behave if it also had conductance X? Patch amplifier Vmem from cell Dynamic clamp: Mathematical Definition of conductance X PC Current command signal to recording Develop method using cultured hippocampal neurones Adding/Subtracting M-current (Kv7) with Dynamic Clamp V Patch clamp amplifier IK(V) XE-991 IKv7 V (current clamp) IKv7 Digitizer ICa(V) ISK(Ca) read V compute df/dt = (f(V)-V)/Kv7 IKv7 = gKv7 × f × (V-VK) write IKv7 Original concept : Sharp et al, 1993 Implementation : Cambridge Conductance (Robinson, 2008) Computer Overview of the neural models: Biological Reality Numerical Simulation • Detailed conductance based models (HH) • Reduced conductance based models (Morris-Lecar) • Two Dimensional Neurons (Fitzhugh-Nagumo) • Integrate-and-Fire Models • Firing rate Neurons (Wilson-Cowan) • Steady-State models • Binary Neurons (Ising) Artificial Analytical Solution